1. Field of the Invention
The present invention relates to an apparatus and methodology for the chromatography of materials, and in particular, chromatography based on molecular size, affinity and the like as used, for example, in the purification, separation or isolation of DNA and RNA fragments, proteins and other molecules.
2. Background Art
Removing unincorporated nucleotides from DNA and RNA fragments, isolating RNA fractions, purifying proteins and other macromolecules, are important procedures having a variety of applications. In DNA and RNA synthesis, unincorporated nucleotides must often be removed when constructing nick-translated probes, RNA probes and end-labeled oligonucleotides, as well as "filled-in" DNA fragments. It is important to separate the unincorporated free-nucleotides from the labeled probe as unincorporated label may bind to the solid support, resulting in unacceptably high levels of background noise. Isolation of RNA fractions may be employed in the separation of, for example, polyadenylated RNA from nonpolyadenylated RNAs. The use of chromatography methods to isolate and identify proteins and other macromolecules is another well known application.
Current chromatography methods, used particularly in connection with DNA and RNA synthesis, include ion-exchange chromatography, several variations of gel chromatography and others. Each has its own disadvantages. For example, ion-exchange methods require a number of steps which may result in a significant investment of time and, in the case of radiolabeled nucleotide filtering, extensive handling of radioactive material. Conventional gel-chromatography "drip" columns are tedious, requiring time to both pour and run. Spin columns, a variation of the "drip" column, are somewhat faster, but risk radiation exposure and contamination in the case of radionucleotide chromatography, and may yield less reliable results.
An alternative chromatography approach which avoids the aforementioned difficulties would therefore be desirable.